Breast cancer development and progression are influenced by steroid hormones, particularly estrogen, via their interaction with specific target cell receptors. Tamoxifen is a non-steroidal antiestrogen which is now the most frequently used drug in breast cancer treatment. Tamoxifen is thought to inhibit breast cancer growth by competitively blocking estrogen receptor (ER), thereby inhibiting estrogen-induced growth. ER, a ligand-dependent transcription factor, meditates most biological effects of estrogens on cell and tumor growth [Katzenellenbogen et al., 2000, Recent Prog Norm Res 55:163–195]. In the adjuvant setting after primary surgery for breast cancer, tamoxifen has been shown to prolong disease-free and overall survival, and it has also been shown to induce remissions in more than half of patients with metastatic disease who have ER-positive tumors [Early Breast Cancer Trials Collaborative Group, 1992, Lancet 339:1–14, 71–85; Saez et al., 1989, Current Clinical Oncology, Alan R Liss Inc., New York, pp. 163–172]. Although tamoxifen is initially effective in many patients, 50% of the patients fail to respond to the drug despite the presence of ER. Furthermore, even patients who initially respond eventually acquire tamoxifen resistance, leading to tumor progression and death. The mechanisms for either intrinsic or acquired tamoxifen resistance are unknown, but they are probably multifactorial.
In both animal models and clinical specimens, lower tamoxifen uptake and somewhat altered tamoxifen metabolism in resistant tumors have been observed, but neither appears to explain tamoxifen stimulation of the resistant tumors. Nor do estrogen receptor losses or mutations appear to explain this phenomenon, although altered expression of transcriptional variant forms of the receptor may well contribute. Pure steroidal antiestrogens such as ICI 182,780 are capable of reversing tamoxifen-stimulated as well as estrogen-stimulated growth of these resistant tumors, and are now in clinical trials for this purpose [Osborne et al., 1994, Breast Cancer Res Treat. 32(1):49–55]. Endocrine treatment of breast cancer is the major form of systemic adjuvant therapy and therapy for metastatic disease. Unfortunately, however, not all tumors will respond, and furthermore, initially responding tumors eventually become resistant to the endocrine treatment, leading to tumor progression and death.
The present inventors are interested in the mechanisms by which tumors develop resistance to tamoxifen, with the ultimate goal of developing new strategies for preventing or reversing the emergence of resistant cells. There are several possible mechanism by which tamoxifen resistance could develop in breast cancer cells. Clues to these mechanisms can be gleaned from an understanding of the myriad effects that tamoxifen has at the cellular level, as outlined in FIG. 1. Tamoxifen binds to the ER and competitively blocks estrogen-induced transcription of specific genes encoding proteins involved with regulation of cell proliferation. Some of these proteins are in fact polypeptide growth factors, such as transforming growth factor α, insulin-like growth factor II, and members of the fibroblast growth factor family, which by autocrine and paracrine mechanisms may enhance tumor growth. Down-regulation of the gene expression of these growth factors by tamoxifen may result in suppression of tumor growth. Oddly enough, breast cancer cells, as well as other tumor cells, may also synthesize and secrete growth inhibitors, such as transforming growth factor-β. Expression of TGFβ is reduced by estrogen, but enhanced by tamoxifen treatment. Thus, increased expression of growth inhibitors by tamoxifen may also contribute to tumor growth suppression. Clearly, alterations in the expression of these growth factors or growth inhibitors, or their specific cell membrane receptors, could provide the tumor cell with sufficient growth stimulation to overcome the tamoxifen block, resulting in tamoxifen resistance. Cross-talk between polypeptide growth factor pathways and ER-mediated events could also theoretically result in tamoxifen resistance. It has been shown, for example, that increasing the level of cellular cyclic AMP pharmacologically alters the cellular response to tamoxifen, converting it from an antiestrogen to a weak estrogen agonist [Fujimoto et al., 1994, Mol Endocrinol. 8(3):296–304]. The mechanism for this phenomenon is not yet understood, but it could be related to changes in the phosphorylation state of the ER itself and/or its coregulatory protein.
Other potential mechanisms for the development of tamoxifen resistance include the loss of or mutations in the ER, or altered expression of the accessory proteins that could modify the transcriptional signal generated by the ligands binding to estrogen receptor. Also, since certain metabolites of tamoxifen are known to be less antiestrogenic, or even to be full estrogen agonists, changed systemic metabolism of tamoxifen or altered uptake or metabolism of tamoxifen in the tumor itself could also result in tamoxifen resistance. Finally, high levels of the so-called antiestrogen binding sites, cytoplasmic binding sites whose function is not yet well understood, could theoretically serve as a sump, soaking up tamoxifen molecules and preventing their binding to ER. Studies of several of these possibilities have been initiated in laboratory models.
Clinical studies with tamoxifen provide several clues for mechanisms by which acquired resistance may develop. Patients whose tumors initially lack ER have a very low response rate to the drug, and thus, selection of an ER-negative clone of tumor cells could result in an estrogen-independent tumor refractory to tamoxifen. Some patients with tamoxifen resistance do develop resistance to all forms of endocrine therapy via selection of an ER-negative tumor cell clone. However, it was recently reported that a series of patients with acquired tamoxifen resistance in whom tumor estrogen and progesterone receptors were measured by both ligand binding and immunohistochemical assays (to circumvent the problem of receptor occupancy by the drug) [Encarnacion et al., 1993, Breast Cancer Res. Treat. 26(3):237–246]. More than 60% of tumors continued to express ER and/or PgR even while progressing in the face of tamoxifen. These data indicate that while ER negativity may account for some cases of resistance, mechanisms of resistance other than receptor loss must be common.
If patients' tumors remain ER-positive after development of tamoxifen resistance, one might expect that some of these tumors have retained estrogen sensitivity and will respond to other endocrine treatments. In fact, clinical experience demonstrates that patients who have initially responded to tamoxifen but who later develop tumor progression, frequently respond to second or third-line endocrine therapies. Thus, acquired tamoxifen resistance in these patients does not necessarily indicate global hormonal unresponsiveness, but rather selective resistance to tamoxifen itself. Although it has not been studied systematically, anecdotal experience suggests that some patients with tamoxifen resistance will respond to a rechallenge with the drug after an interval in which they receive other treatments. Furthermore, clinical reports suggest that patients who receive tamoxifen adjuvant therapy and then later recur, not infrequently will respond to a rechallenge with the drug. This suggests that tamoxifen resistance, in some cases, may not be a permanent phenotype, but rather may be reversible when administration of the drug is stopped. Patients may also respond to an increase in the tamoxifen dose after developing progression with a lower dose schedule. Finally, similar to reports of patients treated with high dose estrogen therapy, some patients who have responded to tamoxifen will have a withdrawal response when the drug is stopped at the time of tumor progression. The prolonged half-life of tamoxifen makes it difficult for clinicians to withhold alternative therapy while waiting for a withdrawal response to the drug. Nevertheless, these data strongly suggest that in some patients with acquired resistance, tamoxifen may actually be stimulating tumor growth.
Two previously published clinical trials also suggest that tamoxifen-stimulated tumor growth may be a cause of tamoxifen resistance in some patients [Pritchard et al., 1980, Cancer Treat Rep. 64(6–7):787–96][Hoogstraten et al., 1984, Cancer 54(10):2248–2256]. In these studies, premenopausal women with advanced breast cancer were treated with second-line ovarian ablation after they first responded and then progressed on tamoxifen. In one of these studies, the secondary response to ovarian ablation was common in patients who had previously responded to tamoxifen, suggesting that tamoxifen treatment served as an in vivo tumor estrogen sensitivity assay. However, in the other study, opposite results were obtained and no patients responded to second-line ovarian ablation. In this latter study tamoxifen therapy was continued after the surgery, while in the first study tamoxifen treatment was stopped. Secondary response to ovarian ablation would not be expected in the latter study if tamoxifen itself was behaving as an estrogen agonist and stimulating tumor growth. Tamoxifen-stimulated tumor growth as a mechanism for acquired resistance is further supported by data from the present inventor's laboratory as well as others using experimental models. A major focus of the present inventor's group is to better understand mechanisms by which tamoxifen-stimulated tumor growth occurs.